The study of signaling pathways in biological cells may lead to greater understanding of cellular responses to various disease-causing agents, resulting in improved treatment alternatives in the form of drugs that target specific pathways. Fluorescence-based biological assays have demonstrated tremendous value in allowing researchers to understand complex biological processes. For example, organic fluorescent molecules or green fluorescent proteins (GFPs) are commonly used materials for fluorescent tagging of biological substances, including cells, and have been utilized in cell imaging, cell tracking, and in vivo monitoring of cellular events. GFPs have excellent biological compatibility because they are genetically encoded and expressed by the cell itself. However, the poor photostability of some organic fluorophores and GFPs may cause difficulties in long-term monitoring of cellular events, where high sensitivity and high image resolution are often desired. Additionally, GFPs require a time-consuming process for establishing stable-expressing clones, which may limit their use in long-term imaging of live cells.
Fluorescent quantum dots (QDs) are nanometer-sized heavy metal or semiconductor particles that can be covalently linked to bio-recognition molecules such as peptides, antibodies, nucleic acids, or small-molecule ligands for application as fluorescent probes. QDs have high absorption coefficients, high photobleaching thresholds, and high quantum yields. However, despite the advantageous photophysical nature of QDs, they are also very highly toxic. To reduce the inherent toxicity of heavy metals, various chemical approaches have been applied, including coordination of small molecules, silylation, and encapsulation of lipids. Cell survivability in particular cell lines has been reported by decorations or biomolecule encapsulation of QDs; however, their toxicity still remains a problem. Initial investigation into CdSe QD toxicity using primary hepatocytes rich in metallothionein as a model for the liver clearly displayed that, under certain conditions, the QDs were cytotoxic. The toxicity was attributed to liberation of Cd2+ ions during oxidation. Also, the surface modification of QDs resulted in decreased emission efficiency and poor colloidal stability. Another side effect of the surface coatings of QDs may be that the coatings can change dramatically with the movement, retention, and distribution of QDs. Also, size may often be the basic determinant for the photophysical properties of quantum dots, wherein a larger QD may have superior optical properties relative to a smaller QD.
Accordingly, improved methods are needed.